The primary focus of this renewal proposal is on how repair enzymes process specific exocyclic DNA adducts, which are formed by diverse environmental mutagens/carcinogens including vinyl compounds, glycidyl ethers, chlorohydroxyfuranones, benzene and therapeutic nitrosoureas such as BCNU or formed by endogenous processes. Unrepaired adducts can block replication or cause mutation since many of these exocyclic adducts are miscoding lesions. The exocyclic adducts to be studied are structurally related but with differing features. Our central hypothesis is that such ring structural features, formed by different carcinogens, will determine specific enzymatic recognition and the type of repair. On this basis, we hope to identify and characterize new repair activities or novel repair enzymes. The proposed approaches from biochemistry and structural studies attempt to define specific structural features or rules that are essential for enzymatic recognition or repair efficiency. In addition, protein-protein interactions in glycosylase excision of exocyclic adducts will be investigated. By using in vitro biochemical approaches and specific oligonucleotides containing a single site-directed adduct, we wish to carry out the following specific aims: (1) To identify DNA glycosylases/endonucleases acting on newly synthesized exocyclic adducts; (2) To examine the initial recognition by mismatch repair (MMR) pathway of the available exocyclic adducts as these lesions form similar structures to mismatches; (3) To explore the potential pathway for p-benzoquinone adduct repair using an in vitro substitution methodology; (4) To study how and to what extent the DNA glycosylases excising exocyclic adducts interact with other cellular proteins such as 5'AP endonucleases, XPG protein and the MMR binding proteins; and (5) Structural studies, such as molecular modeling of damaged DNA/glycosylase complexes, will be performed to aid in understanding how repair enzymes interact with such adducted DNA.